This laboratory has used forward genetic screens and gene expression analysis involving in situ hybridization and DNA microarray technology to identify genes that function in embryonic development. We have identified a mutation of the gene encoding the Mediator component MED12, also called TRAP230. Mediator is a complex of many proteins that mediates the transcriptional effect of sequence-specific factors such as nuclear hormone receptors or homeodomain proteins to RNA polymerase and associated factors. The mutation named kohtalo (kto) affects the development of multiple systems in zebrafish embryogenesis, in particular the brain, neural crest derivatives, and the kidney. Tissue arrangements and morphogenetic movements are particularly affected in the mutation. In particular, the kto gene is required for normal organization of rhombomere boundaries which is critical for proper regionalization of the hindbrain. The mechanisms underlying these processes are currently under study. A different mutant that was characterized affects BAP28, a component of the ribosome biosynthesis pathway. We concluded that the imbalance in ribosome biogenesis in the mutant triggers apoptotic cell death in the embryo by a process that involves the p53 tumor suppressor gene.[unreadable] Zebrafish Protocadherin18 was isolated on the basis of gene expression analysis. We have shown that this protein is found at the cell membrane in multiple embryonic tissues, and that it has a role in cell interactions during morphogenesis in embryonic development. The zebrafish barx1 gene has been isolated and shown to be involved in the regulation of chondrogenesis during pharyngeal arch development. The function of barx1 is required for the normal formation of the craniofacial skeleton. [unreadable] A collaborative project has been carried out to characterize the enzymes responsible for heme biosynthesis in zebrafish and the genes that encode these enzymes. Additional genes involved in hematopoiesis in the zebrafish embryo have been identified and are currently under study.[unreadable] An extensive DNA microarray analysis was carried out to characterize the RNA populations in the zebrafish pineal gland and compare them to the RNA populations in the brain at different stages of development and adult life. The pineal gland has an important role in regulating circadian rhythm in the animal. The pineal transcriptome was analyzed at three embryonic and two adult stages of development, allowing developmental analysis of gene expression in addition to the identification of genes preferentially expressed in the pineal gland. Global analysis of the data by bioinformatic means is currently under way. In addition, individual genes differentially expressed in the pineal gland are being subjected to functional analysis.